GRSDB is a database of G-quadruplexes and contains information on composition and distribution of putative Quadruplex-forming G-Rich Sequences (QGRS) mapped in the eukaryotic pre-mRNA sequences, including those that are alternatively processed (alternatively spliced or alternatively polyadenylated). The data stored in the GRSDB is based on computational analysis of NCBI Entrez Gene entries and their corresponding annotated genomic nucleotide sequences of RefSeq/GenBank.

A database of quadruplex motifs. It is composed of two parts (EuQuad and ProQuad). EuQuad gives information on quadruplex motifs present in human, chimpanzee, rat and mouse genes. ProQuad contains quadruplex information of 146 prokaryotes. Apart from gene-specific searches QuadBase has a number of other modules. ‘Orthologs Analysis' queries for conserved motifs across species in a user-defined manner; ‘Pattern Search' can be used to fetch specific motifs of interest and the ‘Pattern Finder' tool can search for motifs in any given sequence

G-quadruplexes are non-canonical nucleic-acid structures that control transcription, replication, and recombination in organisms. G-quadruplexes are present in eukaryotes, prokaryotes, and viruses. In the latter, mounting evidence indicates their key biological activity. Since data on viruses are scattered, we here present a comprehensive analysis of potential quadruplex-forming sequences (PQS) in the genome of all known viruses that can infect humans. We show that occurrence and location of PQSs are features characteristic of each virus class and family. Our statistical analysis proves that their presence within the viral genome is orderly arranged, as indicated by the possibility to correctly assign up to two-thirds of viruses to their exact class based on the PQS classification. For each virus we provide: i) the list of all PQS present in the genome (positive and negative strands), ii) their position in the viral genome, iii) the degree of conservation among strains of each PQS in its genome context, iv) the statistical significance of PQS abundance. This information is accessible from a database to allow the easy navigation of the results: http://www.medcomp.medicina.unipd.it/main_site/doku.php?id=g4virus. The availability of these data will greatly expedite research on G-quadruplex in viruses, with the possibility to accelerate finding therapeutic opportunities to numerous and some fearsome human diseases.

A database listing potential G-quadruplex regulated genes. G-rich DNA sequences can form G-quadruplexes, a four-stranded structure that is stabilized by planar arrays of four guanines associated with hydrogen bonds. Promoter G-quadruplexes have emerged as a new way to regulate gene transcription, such as in c-MYC expression. Further, G-quadruplex motifs are highly enriched in gene promoter regions in humans and other mammals. Greglist contains genes whose promoter regions have G-quadruplex motifs, and these genes are highly likely to be regulated by G-quadruplexes.

Aptamer is a nucleic acid ligand which specifically binds to its target molecule. Previously, we have designed an identification method of aptamer called “G-quadruplex (G4) promoter-derived aptamer selection (G4PAS)” [1]. In G4PAS procedure, putative G4 forming sequences (PQS) were explored in a promoter region of a target protein in human gene through computational analysis, and evaluated binding ability towards the gene product encoded in the downstream of the promoter. We investigated the topology of the obtained PQSs by circular dichroism measurement, as well as their binding ability against its target protein by surface plasmon resonance measurement and gel-shift assay. Additionally, the presence of nuclear localization signal in the target protein was predicted in silico. This data set summarized all the PQS sequences, these biochemical characteristics, and the presence of nuclear localization signal to address the possibility of binding of these PQS region to the target proteins in vivo. hese data should contribute to increase the success rate of G4PAS, and potentially beneficial for the cell biology field as G4 is supposed to participate in vivo gene regulation.

The physiological role(s) played by G-quadruplexes renders these ‘non-canonical’ DNA secondary structures interesting new targets for therapeutic intervention. In particular, the search for ligands for selective recognition and stabilization of G-quadruplex arrangements has led to a number of novel targeted agents. An interesting approach is represented by the use of metal-complexes, their binding to DNA being modulated by ligand and metal ion nature, and by complex stoichiometry. In this work we characterized thermodynamically and stereochemically the interactions of a Ni(II) bis-phenanthroline derivative with telomeric G-quadruplex sequences using calorimetric, chiroptical and NMR techniques. We employed three strictly related sequences based on the human telomeric repeat, namely Tel22, Tel26 and wtTel26, which assume distinct conformations in potassium containing solutions. We were able to monitor specific enthalpy/entropy changes according to the structural features of the target telomeric sequence and to dissect the binding process into distinct events. Interestingly, temperature effects turned out to be prominent both in terms of binding stoichiometry and ΔH/ΔS contributions, while the final G-quadruplex-metal complex architecture tended to merge for the examined sequences. These results underline the critical choice of experimental conditions and DNA sequence for practical use of thermodynamic data in the rational development of effective G-quadruplex binders.

G-quadruplex structures in the 5’ UTR of mRNAs are widely considered to suppress translation without affecting transcription. The current study describes the comprehensive analysis of proteins binding to four different G-quadruplex motifs located in mRNAs of the cancer-related genes Bcl-2, NRAS, MMP16, and ARPC2. Following metabolic labeling (Stable Isotope Labeling with Amino acids in Cell culture, SILAC) of proteins in the human cell line HEK293, G-quadruplex binding proteins were enriched by pull-down assays and identified by LC-orbitrap mass spectrometry. We found different patterns of interactions for the G-quadruplex motifs under investigation. While the G-quadruplexes in the mRNAs of NRAS and MMP16 specifically interacted with a small number of proteins, the Bcl-2 and ARPC2 G-quadruplexes exhibited a broad range of proteinaceous interaction partners with 99 and 82 candidate proteins identified in at least two replicates, respectively. Among the interaction partners were many proteins known to bind to RNA, including multiple heterogenous nuclear ribonucleoproteins (hnRNPs). The identified ribosomal proteins are likely to reflect stalling of the ribosome by RNA G-quadruplex structures. In addition, several proteins were identified that have not previously been described to interact with RNA. Gene ontology analysis of the set of candidate proteins revealed that many interaction partners are known to be tumor related. The majority of the identified RNA G-quadruplex interacting proteins are thought to be involved in post-transcriptional processes, particularly in splicing. These findings indicate that protein-G-quadruplex interactions may be relevant to the regulation of mRNA maturation and may play an important role in tumor biology.

This is the NMR dataset for the manuscript 'NMR screen reveals the diverse structural landscape of a G-quadruplex library'

Abstract

G-quadruplexes are noncanonical nucleic acid structuresformed by stacked guanosine tetrads. Despite their functional andstructural diversity, a single consensus model is typically used todescribe sequences with the potential to form G-quadruplexstructures. We are interested in developing more specific sequencemodels for G-quadruplexes. In previous work, we functionallycharacterized each sequence in a 496-member library of variants of amonomeric reference G-quadruplex for the ability to bind GTP,promote a model peroxidase reaction, generate intrinsic fluorescence,and to form multimers. Here we used NMR to obtain a broad overviewof the structural features of this library. After determining the 1H NMRspectrum of each of these 496 sequences, spectra were sorted intomultiple classes, most of which could be rationalized based onmutational patterns in the primary sequence. A more detailed screenusing representative sequences provided additional information aboutspectral classes, and confirmed that the classes determined based onanalysis of 1H NMR spectra are correlated with functional categoriesidentified in previous studies. These results provide new insights intothe surprising structural diversity of this library. They also show howNMR can be used to identify classes of sequences with distinctmutational signatures and functions.

Link to journal article: https://doi.org/10.1002/chem.202401437

The comprehensive list of lncRNAs dysregulated in diverse human cancers, their expression patterns, methodologies of their identification, and references to research articles (PubMed ID) used to gather these details were obtained from the Lnc2Cancer 3.0 dataset. The aliases of all these lncRNAs were manually compiled from the GeneCards dataset. The nucleotide sequences and the corresponding NCBI accession numbers of all the identified lncRNAs, including their functional transcript variants, with “validated” or “reviewed” RefSeq status, were retrieved from NCBI Nucleotide.For identification of Putative Quadruplex-forming Sequences (PQS) within these lncRNAs, their FASTA sequences are imported to the QGRS mapper, a tool that presents data on the constitution and distribution of Quadruplex-forming G-rich sequences (QGRS). The PQS within these lncRNAs are also identified using G4Hunter, a tool for identifying putative G4-forming motifs based on the G-richness and G-skewness of the query sequence.The details of experimentally-validated RNA and protein interacting partners of catalogued lncRNAs were sourced from NPInter v4.0 and LncTarD 2.0 datasets. The data obtained from LncTarD 2.0 was filtered to present the data exclusively in the context of human cancers. The information on the experimentally-validated RNA G4-binding proteins (RGBPs) interacting with the catalogued lncRNAs was obtained from QUADRatlas and G4IPDB datasets, supplemented by scientific literature mining.

The spectral properties of meso-tetrakis (N-methylpyridinium-4-yl)porphyrin (TMPyP) in the presence of parallel and antiparallel G-quadruplexes formed from a thrombin-binding aptamer G-quadruplex (5′-G3T2G3TGTG3T2G3) were investigated in this study. Red shift and hypochromism in the Soret absorption band of TMPyP were observed after binding to both parallel and antiparallel G-quadruplexes. The extent of changes in the absorption spectra were similar for both conformers. No circular dichroism spectrum was induced in the Soret region for both parallel and antiparallel G-quadruplexes. This is suggest that there is no or very weak interaction between electric transitions of nucleobases and porphyrin molecule. The accessibility of the neutral quencher I2 to the G-quadruplex-bound TMPyP was similar for both parallel and antiparallel G-quadruplexes. All these observations suggest that TMPyP was bound at the outside of the quadruplexes, and conceivably interacted with the phosphate group via a weak electrostatic interaction. Communicated by Ramaswamy H. Sarma

Biological Magnetic Resonance Bank Entry 16356: Structure of a two-G-tetrad intramolecular G-quadruplex formed by a variant human telomeric sequence in K+ solution

G-quadruplex (G4) structures can form in guanine-rich DNA or RNA and have been found to modulate cellular processes including replication, transcription, and translation. Many studies on the cellular roles of G4s have focused on eukaryotic systems, with far fewer probing bacterial G4s. Using a chemical-genetic approach, we identified genes in Escherichia coli that are important for growth in G4-stabilizing conditions. Reducing levels of elongation factor Tu or slowing translation elongation with chloramphenicol suppress the effects of G4 stabilization. In contrast, reducing the expression of certain translation termination or ribosome recycling proteins is detrimental to growth in G4-stabilizing conditions. Proteomic and transcriptomic analyses demonstrate that ribosome assembly factors and other proteins involved in translation are less abundant in G4-stabilizing conditions. Our results suggest that RNA G4s can present barriers to E. coli growth and reducing the rate of translatio..., Strain construction All cells used in this study are derived from an Escherichia coli MG1655 parent strain unless otherwise specified. For CRISPR interference strains, strains were a gift from Jason Peters (46). To generate E. coli knockout strains, P1 transductions were carried out using Keio collection strains as the donor strain (63,64). P1 phage lysate was grown on Keio collection donor strains, which was used to transduce the MG1655 strains or CRISPR interference strains (to make the tolC knockout of selected CRISPRi strains) which were sensitive to kanamycin. To validate strains, transductions were grown on LB plates supplemented with 50 µg/mL kanamycin and screened using colony PCR to validate proper insertion of the kanamycin resistance cassette. To remove the kanamycin resistant cassette from MG1655 tolC::kan to enable additional P1 transductions in this strain, MG1655 tolC::kan electrocompetent cells were generated and transformed with a plasmid encoding the FLP recombinase (p..., , # Altering translation allows E. coli to overcome chemically stabilized G-quadruplexes

https://doi.org/10.5061/dryad.zcrjdfnn9

Description of the data and file structure

The data included in this Dryad submission was collected in order to understand how the model organism* Escherichia coli*Â overcomes stabilized G-quadruplexes. This work involved a multi-omics approach to studying how the G-quadruplex stabilizers NMM and Braco-19 impact growth, gene importance, and mRNA/proteomic abundance in G-quadruplex stabilizing conditions.Â

Files and variables

File: Western_total_protein_stain_1.tif

Description: Total protein staining for western blots assessing EF-Tu abundance in ∆tolC, ∆tolC tufA::kan, and ∆tolC tufB::kan cells. Order of gel is ∆tolC, ∆tolC tufA::kan, and ∆tolC tufB::kan at three different dilutions added into the gel. This is the total protein stain image used in the figure of the manuscript.

File: Wester...

G-quadruplex-forming small RNA inhibits coronavirus and influenza A virus replication

https://doi.org/10.5061/dryad.8931zcs1s

Principle Investigator Contact Information

Name: Shusuke Kuge

Institution: Division of Microbiology, Faculty of Pharmaceutical Sciences, Tohoku Medical and Pharmaceutical University.

e-mail: skuge@tohoku-mpu.ac.jp

QUMA-1 stain for the cells

A549 cells were cultured in 96-well plates (ViewPlate). After being washed with MEM, cells were cultured in the presence of 2 μM of each PS-RNA for 17 h. Cells were then washed with MEM and fixed with 4% paraformaldehyde in PBS for 15 min on ice. After washing three times with PBS, the cells were observed after 7 h or later by adding PBS containing 50 nM QUMA-1 and 1 μg/mL DAPI. Confocal imaging was performed using LSM900 (Zeiss) equipped with Airyscan 2 using Plan-Apochromat 63x/1.40 Oil DIC M27 (objective). Excitation and detection wa...

ENCODES a protein that exhibits G-quadruplex DNA binding (ortholog); single-stranded DNA binding (ortholog); single-stranded RNA binding (ortholog); INVOLVED IN G-quadruplex DNA formation (ortholog); negative regulation of transcription by RNA polymerase II (ortholog); positive regulation of cell population proliferation (ortholog); ASSOCIATED WITH genetic disease (ortholog); myotonic disease (ortholog); myotonic dystrophy type 1 (ortholog); FOUND IN cytosol (ortholog)

Conformational changes in DNA G-quadruplex (GQ)-forming regions affect genome function and, thus, compose an interesting research topic. Computer modelling may yield insight into quadruplex folding and rearrangement, particularly molecular dynamics simulations. Here, we show that specific parameters, which are distinct from those commonly used in DNA conformational analyses, must be introduced for adequate interpretation and, most importantly, convenient visual representation of the quadruplex modelling results. We report a set of parameters that comprehensively and systematically describe GQ geometry in dynamics. The parameters include those related to quartet planarity, quadruplex twist, and quartet stacking; they are used to quantitatively characterise various types of quadruplexes and rearrangements, such as quartet distortion/disruption or deviation/bulging of a single nucleotide from the quartet plane. Our approach to describing conformational changes in quadruplexes using the new parameters is exemplified by telomeric quadruplex rearrangement, and the benefits of applying this approach to analyse other structures are discussed.

Inhibition of telomerase by inducing/stabilizing G-quadruplex formation is a promising strategy to design new anticancer drugs. We synthesized and characterized a new dinuclear complex [(dmb)2Ru(obip)Ru(dmb)2]4+ (dmb = 4,4’-dimethyl-2,2’-bipyridine, obip = (2-(2-pyridyl)imidazo[4,5-f][1,10]phenanthroline) with high affinity for both antiparallel and mixed parallel / antiparallel G-quadruplex DNA. This complex can promote the formation and stabilize G-quadruplex DNA. Dialysis and TRAP experiments indicated that [(dmb)2Ru(obip)Ru(dmb)2]4+ acted as an excellent telomerase inhibitor due to its obvious selectivity for G-quadruplex DNA rather than double stranded DNA. In vitro co-culture experiments implied that [(dmb)2Ru(obip)Ru(dmb)2]4+ inhibited telomerase activity and hindered cancer cell proliferation without side effects to normal fibroblast cells. TUNEL assay indicated that inhibition of telomerase activity induced DNA cleavage further apoptosis in cancer cells. Therefore, RuII complex represents an exciting opportunity for anticancer drug design by specifically targeting cancer cell G-quadruplexes DNA.

Molecular Dynamics simulation trajectories of basket-type DNA G-quadruplex (PDB: 2KF7). The structures have been simulated in the temperatures 300 K and 360 K in pure water, 1 M TMAO solution, 2 M urea solution and the mixture of 1 M TMAO and 2 M urea.

Biological Magnetic Resonance Bank Entry 34443: Timeless couples G quadruplex detection with processing by DDX11 during DNA replication

G-quadruplex, assembled from a square of guanine (G) molecules, is an important structure with crucial biological roles in vivo but also a vesitile template for ordered functional material. Although the understanding of G-quadruplex structures is the focus of numerous studies, little is known regarding the control of G-quartet stacking modes and the spontaneous orientation of G-quadruplex fibrils. Here, the effects of different metal ions and their concentrations on stacking modes of G-quartets are elucidated. Monovalent cations (typically K+) facilitate the formation of G-quadruplex hydrogels with both heteropolar and homopolar stacking modes, showing weak mechanical strength. In contrast, divalent metal ions (Ca2+, Sr2+, and Ba2+) at given concentrations can control G-quartet stacking modes and increase the mechanical rigidity of the resulting hydrogels through ionic bridge effects between divalent ions and borate. We show that for Ca2+ and Ba2+ at suitable concentrations, the assembl...

G-quadruplex (or G4) structures are non-canonical DNA structures that form in guanine-rich sequences and threaten genome stability when not properly resolved. G4 unwinding occurs during S phase via an unknown mechanism. Using Xenopus egg extracts, we define a three-step G4 unwinding mechanism that acts during DNA replication. First, the replicative helicase (CMG) stalls at a leading strand G4 structure. Second, the DHX36 helicase mediates the bypass of the CMG past the intact G4 structure, which allows approach of the leading strand to the G4. Third, G4 structure unwinding by the FANCJ helicase enables the DNA polymerase to synthesize past the G4 motif. A G4 on the lagging strand template does not stall CMG, but still requires active DNA replication for unwinding. DHX36 and FANCJ have partially redundant roles, conferring robustness to this pathway. Our data reveal a novel genome maintenance pathway that promotes faithful G4 replication thereby avoiding genome instability.